摘要
根据柑橘黄龙病(citrus Huanglongbing,HLB)病菌16SrDNA、16S/23S核糖体基因间隔区(ribosomal intergenic region,RIR)、核糖体蛋白β操纵子(β-operon)和外膜蛋白(out membrane protein,OMP)基因序列设计了8对引物,通过常规和巢式PCR方法对各引物的检测灵敏度进行了比较。结果表明,不同引物的检测灵敏度不同。对于上述任何基因,巢式PCR的灵敏度均比常规PCR至少高103倍;对于同一种基因,扩增短片段的引物比扩增长片段的引物灵敏度高或相当;在扩增产物大小相同时,用以扩增核糖体蛋白β操纵子基因的引物较其他三种稍高。对不同症状柑橘样品的检测进一步验证了该结果。因此,对于柑橘黄龙病的检测,可优先考虑使用巢式PCR方法,若使用常规PCR,则宜选用具有高灵敏度的扩增小片段引物。
Eight primer pairs were designed according to gene sequences of 16S rDNA,16S/23S ribosomal intergenic region,ribosomal protein β-operon and out membrane protein of citrus Huanglongbing pathogen,and then the detection sensitivity of different primer pairs were compared by conventional PCR and nested PCR.The result showed that different primer pairs had distinct detection sensitivity.For any gene,the sensitivity of nested PCR was more than 103 times higher than that of conventional PCR.For the same gene,the sensitivity of primers for amplifying short fragments were higher than or equal to that of primers for amplifying long fragments.When the size of amplified product was the same,the primers for amplifying ribosomal protein gene had slightly higher sensitivity than the other three ones.The detection for citrus samples with different symptoms further verified this result.Therefore,nested PCR should be preferred to detect citrus Huanglongbing.If using conventional PCR,the primers with high sensitivity for amplifying small fragments should be chosen
出处
《热带作物学报》
CSCD
2010年第8期1280-1287,共8页
Chinese Journal of Tropical Crops
基金
国家现代柑橘产业技术体系华东柑橘综合试验站专项经费(农业部农科教发[2007]14号文件)
浙江省农科院科技创新能力提升工程项目(No.2009R27Y01D01)
台州市科技计划项目(No.091KY03)资助
